The term heavy metal is used for any metallic element that has a relatively high density and is toxic at low concentrations [20]. They cause adverse effects on plant physiology such as heavy metal toxicity, inhibition of respiration and photosynthesis, changes in the plant-water relationship, decreased plasma membrane permeability in stem cells and negative effects on the metabolic activities of enzymes [21,20]. Since cadmium is used extensively in industry, it can contaminate foods with air and water [17]. Lead mostly emerges as a result of the combustion of gasoline used in motor vehicles and contaminates sea creatures as a result of transporting industrial wastes with water [17]. High concentrations of lead, which is easily taken from the soil and accumulated in different organs of plants, cause stress, leading to effects such as reduction in root elongation and biomass in the plant, inhibition in chlorophyll biosynthesis, triggering or inhibition of some enzyme activities [22,23]. (Fargasova, 1994; Miranda and Ilangovan, 1996). The responses of plants to lead stress may vary depending on the amount present and the genotype of the plant subjected to the stress. The selectivity of plant species, even genotypes of the same species, in terms of lead uptake, their accumulation capacity and tolerance to lead toxicity may differ [24,25,26]. Nickel contamination occurs in the hydrogenation of vegetable oils. While most of the nickel taken orally is excreted by the body, some of it can accumulate in tissues such as lungs, intestines and skin [17]. The amount of land suitable for agriculture in the world is very low. Heavy metal pollution means lost soils [17]. The lead and copper concentrations in the soil have decreased half in 740–5900 and 310–1500 years, respectively [27,28].
There are many studies on the responses and adaptations of plants to heavy metals. Mercury used in many areas of the industry is not necessary for plants and even low concentrations can cause serious toxicity [29,30]. It also causes plant growth to slow down [30]. In a study carried out by [30], the effects of mercury at different concentrations (0.015625, 0.03125, 0.0625, 0.125, 0.250, 0.500, 1.000 ve 2.000 mM) on mitosis division of root cells were investigated in garlic (Allium sativum). Although a significant difference was not found in germination at low concentrations, decreasing was observed at high concentrations and root development was completely inhibited above 0.250 mM. At all concentrations, root growth was prevented according to the control. With the dose increase, it was determined that cell division decreased and some mitotic abnormalities increased. It is stated that garlic is not a tolerant plant for mercury pollution. It was aimed to reveal the effect of humic acid applications on some morphological and biochemical properties of lettuce grown under heavy metal stress by [31]. In the study carried out under controlled greenhouse conditions, 4 different humic acid doses (0, 2, 4, 8 L da− 1) and 4 different irrigation waters (Control: 0 ppm; I. Mixture: 0.2 ppm Cu + 0.01 ppm Cd + 5 ppm Pb + 2 ppm Zn, Mix II: 0.4 ppm Cu + 0.02 ppm Cd + 10 ppm Pb + 4 ppm Zn, Mixture III: 0.8 ppm Cu + 0.04 ppm Cd + 20 ppm Pb + 8 ppm Zn) were used. The plants were irrigated for 4 weeks with with the specified contents and at the end of this period. In the study; shoot dry and fresh weight, shoot and root length, leaf area, MDA amount, superoxide dismutase and glutathione reductase enzyme activities were investigated. The highest toxic effect was found in the mix III. MDA amounts and antioxidative enzyme activities increased in plants irrigated with water containing heavy metal mixture. 4 L/da humic acid dose was found to be more stable and effective than other doses in reducing the negative effects of heavy metal stress. Humic acid applications were effective on reducing the negative effects of heavy metal stress on growth and development. Tolerance of eggplant rootstock breeding genotypes to lead (Pb) stress was investigated by [26]. In the research, 6 commercial eggplant rootstocks and 4 local eggplant genotypes were used. Two local genotypes having high tolerance to salt and drought stress, commercial rootstocks and other local genotypes were compared. 0, 150 and 300 ppm Pb solutions were applied to eggplant seedlings at the 4–5 true leaves stage. Twenty days after the application, shoot fresh and dry weight, root fresh and dry weight, shoot and root length, leaf area were determined. Köksal F1 and AGR703 commercial rootstocks had higher Pb tolerance. However, positive results were obtained in terms of some parameters from local 2 genotypes having high abiotic stress tolerance. These two genotypes showed almost the same resistance as Köksal F1 and AGR 703 rootstocks at 150 ppm Pb dose, however it was observed significant growth losses with the increase in stress dose. It is thought that different response may be obtained when the dose is given in small increments (300 ppm / 20 ppm = 15 times irrigation). Local genotypes sensitive to other stress factors were affected by both low and high doses. It is recommended to carry out the experiments at advanced developmental stages and to control the long-term cumulative density. In a study of [32], the morphological effects and median effective concentration (EC50) values of cadmium (Cd), chromium (Cr) and lead (Pb) on Ceratophylllum demersum L. and Pogostemon erectus (Dalzell) Kuntze were investigated. In terms of EC50 values, the most toxic heavy metals for C. demersum and P. erectus were recorded as Cd > Cr > Pb. High levels of heavy metals increased the toxic effect on the plant. In general, chlorosis, leaf loss, softening and deaths were observed in plants under metal toxicity. In a study carried out by [20], the cytogenetic effects of nickel (Ni) and lead (Pb) on Allium sativum (garlic) were investigated. Nickel nitrate - Ni(NO3)2 and lead nitrate - Pb(NO3)2 solutions were used at 50, 150 and 450 ppm concentrations for 72 hours. The results have shown that it has a strong inhibitor effect on the rhizogenesis process associated with the increasing concentration of these heavy metals and caused a significant mitodepressive effect in meristematic cells. At the same time, various chromosomal deviations were recorded. In a study carried out by [28], the tolerances of T. aestivum and P. sativum seedlings to copper (copper sulfate: 0.15, 0.30 and 0.60 g L− 1) and lead (lead nitrate: 0.5, 1.0 and 1.5 g L− 1) were investigated. In each experimental group, 50 seeds were placed on filter paper in five 500 mL transparent plastic containers. 10 mL of heavy metal salt solution (copper sulfate 0.15, 0.30 and 0.60 g L− 1 or lead nitrate 0.5, 1.0 and 1.5 g L− 1) were poured into the containers. The containers were closed with transparent lids and incubated at 20–22°C under 17-hours light conditions. Every two days, the same volume of salt solution was added to the containers in the experimental groups and distilled water was added to the control group. At all toxic concentrations, the germination capacity of T. aestivum was decreased: lead nitrate: 19–38%, copper sulfate: 23–58%. Root length and shoot length also decreased. In P. sativum, lethal concentrations for T. aestivum did not show any effect, except for root length. The possible reason for this is explained in the study as leguminous plants contain more protective peptides (phytochelatins and metallothioneins) that chelate heavy metals in the cytoplasm and transmit heavy metals to the vacuole [33]. In addition, it is presented that legume plants contain more protein that increases resistance to environmental stress factors.
Garlic is one of the valuable foods from the past to the present due to its nutritional and medical importance. The aroma and taste of garlic in Turkish cuisine is also important and therefore the expectations of Turkish consumers from garlic with domestic origin cannot be met with imported garlic [34,35]. One of the important advantages of tissue culture is in vitro selection. Significant advantages such as profit from labor, cost and time can be provided by in vitro selection. Then, the results obtained can be verified in in vivo. Although tissue culture techniques in garlic are generally used for obtaining virus-free plants or mass production [36], garlic is also suitable plant for in vitro selection. Therefore, garlic has been selected in this study and tolerance to some heavy metals has been tested in vitro conditions. There are studies investigating tolerance to different heavy metals in different plant species. Plant tissue culture techniques is a useful tool for obtaining large amounts of biomass for repopulation of areas that are deteriorated due to phytoremediation and the presence of heavy metals [37]. There are several abandoned mining areas around the Mediterranean Basin and gives the opportunity to use the ability of selected plants to use for the repopulation of phyoremedation and abandoned mining areas [37]. Two domestic Mediterranean plants (Dittrichia viscosa (L.) Greuter and Cistus monspeliensis L.) were evaluated for phytoremediation on the Elba island. The selected species have been micropropagated in different nutrient media to find optimum growth conditions. Different combinations have been created to mimic the original soil (excessive acidic pH and the presence of heavy metals) [37]. Growth of Prosopis laevigata seedlings, their survival situation and their Pb (II) and Ni (II) uptake were determined to evaluate bioacumulation capacities by [38]. Seedlings were cultured on modified Murashige and Skoog including 10 g L-1 sucrose, Ni (II) at the doses of 0.63, 1.26, 2.10, 4.20 mM and Pb (II) at the doses of 0.0, 0.23, 0.45, 0.90, 1.50, 3.0 mM for 50 days. Germination occurred at all doses, however smaller plants with less leafy and secondary roots were obtained. When the seedlings were cultivated at 1.26 mM Ni and 3.0 mM Pb, it was observed 2582 and 3 895 mg Ni kg-1 and 27 300 and 40 666 mg Pb kg-1 accumulation. In vitro culture was recommended to estimate the responses of plants to environmental pollutants and to reduce the cost of plant experiments. Tolerance to high cadmium concentrations has been investigated in two species of shrubs adabted to Pb: Alyssum montanum and Daphne jasminea in in vito condition by [39]. 0.5, 2.5 and 5.0 µm CdCl2 were applied to the shoot cultures for 16 weeks. In both species, rooted in vitro plants suitable for acclimatization were obtained in the presence of Cd. In A. montanum, low and moderate Cd dose induced proliferation, rooting and biomass production. The growth tolerance index (GTI) in the shoots applied Cd ranged from 120–215% and it was found to be 51–202% in the roots. In contrast, proliferation and rooting of D. jasminea applied Cd was inhibited and GTI in shoots decreased with the increasing Cd doses. However, the accumulation of biomass of the roots subjected to Cd was higher. Interestingly, D. jasminea has accumulated a higher amount of Cd than A. montanum in the roots and immobilized this metal in the root system. On the contrary, A. montanum transferred part of the accumulated Cd to the shoots. [40] stated that the harmful effects of exposure to heavy metals can be mitigated by some mineral elements such as selenium (Se). In their study, they analyzed the morphophysiological changes of Aechmea blanchetiana in response to co-exposure to Pb and Se in in vitro culture. Plants were cultured in nutrient media containing 0 or 4 µM Se and after 72 days of culture, a biphasic medium was created by adding sterilized solution containing 0, 500, 1000, or 2000 µM Pb. The number and diameter of xylem veins of the leaves decreased with increasing Pb concentrations. Plants grown in Pb medium without Se exhibited stress properties such as lower photosynthetic pigment, magnesium and manganese content, and lower RC/CSM.